Frequent detection of T cells with mutations of the hypoxanthine-guanine phosphoribosyl transferase gene in patients with paroxysmal nocturnal hemoglobinuria.

Acquired mutations of the PIG-A gene result in the hemolysis characteristic of paroxysmal nocturnal hemoglobinuria (PNH). Although the etiology of the mutation(s) is unclear, mutable conditions have been suggested by the coexistence of multiple clones with different mutations of PIG-A and by the appearance of leukemic clones in patients with PNH. This study sought to test this hypothesis by examining the frequency of hypoxanthine-guanine phosphoribosyl transferase (HPRT) gene mutations, identified by both resistance to 6-thioguanine (6-TG) and gene analysis. T-cell colonies resistant to 6-TG formed in methylcellulose culture were found in 8 (67%) of 12 PNH patients and 3 (18%) of 17 age-matched healthy volunteers (P <.02, Fisher exact probability test). The incidence of resistant colonies ranged from 40 to 367 (mean 149, x 10(-7)) in the 8 patients and from 1 to 16 (mean 7, x 10(-7)) in the 3 healthy donors. Thus, the HRPT gene mutated more frequently in patients with PNH than in healthy controls (P <.02, Mann-Whitney test). Analysis of bone marrow cells supported these findings. Like the PIG-A mutations in PNH, the HPRT mutations were widely distributed in the coding regions and consisted primarily of base deletions. Unlike PNH cells, 6-TG-resistant cells expressed CD59, indicating that the HPRT mutations did not occur in PNH clones. No correlation was noted between HPRT mutation frequency and content of therapy received by the patients. It is concluded that in PNH patients, conditions exist that favor the occurrence of diverse somatic mutations in blood cells.     

日本血吸虫尼克酰胺磷酸核糖转移酶的生物信息学分析

目的对日本血吸虫尼克酰胺磷酸核糖转移酶（SjNAMPT）基因序列及蛋白序列进行生物信息学分析,为该基因的克隆表达和开发应用奠定基础。方法利用Internet在线分析程序和相关工具软件分析SjNAMPT基因的开放阅读框,分析基因编码蛋白的理化性质、结构域,并预测其空间结构和功能。结果 SjNAMPT蛋白由179个氨基酸组成,分子量为19.63kDa,理论等电点为7.64,为稳定性蛋白,无跨膜区和螺旋卷曲结构,不含有信号肽,含有多个磷酸化位点,该蛋白属于磷酸核糖转移酶保守结构域家族,亚细胞定位于细胞质,二级结构以无规卷曲为主,SjNAMPT蛋白序列与其他物种同种蛋白序列相似性较低。结论 SjNAMPT蛋白为日本血吸虫特异性蛋白,具有一定酶活性,可能与日本血吸虫嘌呤代谢和糖代谢有关,值得进一步研究。     

A case of acquired FXIII deficiency with severe bleeding symptoms

Acquired factor XIII (FXIII) deficiency due to an autoantibody against FXIII is a very rare, yet potentially life-threatening bleeding disorder. As the standard coagulation tests (prothrombin time and activated partial thromboplastin time) are normal, the specialized tests are required to make an accurate diagnosis. Here, we report a case of acquired FXIII deficiency with severe bleeding symptoms. A 75-year-old man was referred to our hospital because of severe bleeding tendency after a tooth extraction. Laboratory findings showed that routine coagulation studies were normal, but factor XIII (FXIII) activity was low (3%). The presence of FXIII inhibitor was detected with dot blotting studies. Although the bleeding tendency was very severe, it was successfully controlled by infusion of FXIII concentrates combined with immunosuppressive treatment (oral prednisolone). Fibrin cross-linking study showed the significant delay of the γ-chain dimer and α-chain polymer formation. Western blotting revealed the marked decrease in FXIII-A level. The mixing study of FXIII activity measured using amine-incorporation assay showed the incomplete inhibition pattern. There seems to be little agreement as to the treatment strategy of acquired FXIII deficiency. In this patient, the use of FXIII concentrates was very useful in the initial treatment of bleeding symptom. The use of steroids was also effective in increasing FXIII activity without any serious complications.     

Biochemical basis of hypoxanthine-guanine phosphoribosyltransferase deficiency in nine families

Summary The concentration of hypoxanthine-guanine phosphoribosyltransferase (HPRT) cross-reacting material (CRM) was determined in haemolysates and/or lymphoblast lysates from nine patients with complete or partial deficiency of HPRT activity. Two of the patients had the fully developed Lesch-Nyhan syndrome and although they had undetectable HPRT activity, small amounts of CRM were found. HPRT-specific mRNA was not detected in lymphoblast lysates from one of these patients, while lysates from the other had a much reduced concentration. Samples from three patients with <0.1% of normal HPRT activity but with minor or no neurological manifestations were also found to contain small amounts of CRM. The other four patients whose HPRT activities ranged from 3 to 10% of normal were found to have CRM concentrations which varied from 26 to 100% of normal. In one patient with a partial deficiency theK _m for 5-phospho--d-ribosyl-1-pyrophosphate was five times normal.     

Heterozygous carriers of succinyl-CoA:3-oxoacid CoA transferase deficiency can develop severe ketoacidosis

Succinyl-CoA:3-oxoacid CoA transferase (SCOT, gene symbol OXCT1) deficiency is an autosomal recessive disorder in ketone body utilization that results in severe recurrent ketoacidotic episodes in infancy, including neonatal periods. More than 30 patients with this disorder have been reported and to our knowledge, their heterozygous parents and siblings have had no apparent ketoacidotic episodes. Over 5 years (2008–2012), we investigated several patients that presented with severe ketoacidosis and identified a heterozygous OXCT1 mutation in four of these cases (Case1 p.R281C, Case2 p.T435N, Case3 p.W213*, Case4 c.493delG). To confirm their heterozygous state, we performed a multiplex ligation-dependent probe amplification analysis on the OXCT1 gene which excluded the presence of large deletions or insertions in another allele. A sequencing analysis of subcloned full-length SCOT cDNA showed that wild-type cDNA clones were present at reasonable rates to mutant cDNA clones. Over the following 2 years (2013–2014), we analyzed OXCT1 mutations in six more patients presenting with severe ketoacidosis (blood pH ≦7.25 and total ketone body ≧10 mmol/L) with non-specific urinary organic acid profiles. Of these, a heterozygous OXCT1 mutation was found in two cases (Case5 p.G391D, Case6 p.R281C). Moreover, transient expression analysis revealed R281C and T435N mutants to be temperature-sensitive. This characteristic may be important because most patients developed ketoacidosis during infections. Our data indicate that heterozygous carriers of OXCT1 mutations can develop severe ketoacidotic episodes in conjunction with ketogenic stresses.     

维生素B12缺乏致严重体位性低血压一例

体位性低血压是引起晕厥的第二大病因,多见于合并高血压、糖尿病、神经系统退行性变的老年人群,然而其发病隐匿,经常被漏诊、误诊。该文报道1例由维生素B12缺乏导致的严重体位性低血压患者。以期加深临床上对维生素B12缺乏导致体位性低血压的认识。     

A case of severe B cell deficiency after allogeneic stem cell transplantation

Insufficient immunological reconstitution is one of the serious complications of allogeneic stem cell transplantation (SCT). We report a case of severely impaired B-lymphopoiesis after allogeneic SCT for CML. The patient's bone marrow and blood cells display complete chimerism and he is currently free from leukemia. His serum immunoglobulin levels are below detection level, and B cells are absent at 2 years post transplant in both the bone marrow and blood. Other populations appear to be normal. To the best of our knowledge, this is the first report of B-lymphopoiesis being undetectable more than 2 years after allogeneic SCT.     

Xanthine nephropathy during chemotherapy in deficiency of hypoxanthine-guanine phosphoribosyltransferase.

We describe an episode of obstructive uropathy produced by xanthine precipitation in the tubules of the kidney of a patient with histiocytic lymphoma during intensive chemotherapy, despite allopurinol therapy. Urinary oxypurine-uric acid ratio suggested a subclinical deficiency of hypoxanthine-guanine phosphoribosyltransferase. Results of an assay of this enzyme confirmed the abnormality. Both parents and three brothers of the patient had normal enzyme activity. The continued importance of adequate hydration for patients who receive allopurinol during initial periods of cancer therapy is emphasized.     

Erythrocyte phosphoribosylpyrophosphate concentrations in heterozygotes for hypoxanthine-guanine phosphoribosyltransferase deficiency

Heterozygotes for the deficiency of hypoxanthine-guanine phosphoribosyltransferase (HGPRT) demonstrate either normal erythrocyte HGPRT activities in families in which the hemizygote shows the Lesch-Nyhan syndrome, or a range of erythrocyte HGPRT activities between 20% of normal and completely normal values in families in which the mutation results in urate overproduction with few or no neurologic signs. Phosphoribosylpyrophosphate (PRPP) concentrations in erythrocytes from heterozygotes for HGPRT deficiency were shown to correlate inversely with HGPRT activities and directly with APRT activities in erythrocyte lysates. Since no evidence of increased synthesis of PRPP was obtained, these elevated PRPP concentrations can best be attributed to reduced utilization of PRPP in the HGPRT-catalyzed reaction. As shown in hemizygotes, the increased PRPP concentrations in these heterozygotes could result in stabilization of the APRT enzyme and elevation of APRT activities.     

Isolation and characterization of a full-length expressible cDNA for human hypoxanthine phosphoribosyl transferase.

We have cloned a full-length 1.6-kilobase cDNA of a human mRNA coding for hypoxanthine phosphoribosyltransferase (HPRT; IMP:pyrophosphate phosphoribosyltransferase, EC 2.4.2.8) into a simian virus 40-based expression vector and have determined its full nucleotide sequence. The inferred amino acid sequence agrees with a partial amino acid sequence determined for authentic human HPRT protein. Transfection of HPRT-deficient mouse LA9 cells with the purified plasmid leads to the expression of human HPRT enzyme activity in cells stably transfected and selected for enzyme activity in hypoxanthine/aminopterin/thymidine medium.     

Kelley-Seegmiller syndrome in a patient with complete hypoxanthine-guanine phosphoribosyltransferase deficiency

Different degrees of hypoxanthine guanine phosphoribosyltransferase (HPRT) deficiency are associated with hyperuricemia, uric acid nephrolithiasis and severe gout. Up to 25-30% of HPRT deficient patients, indicated as neurological variants or HPRT-related hyperuricemia with neurological dysfunction (HRND), may develop neurological manifestation, from mild to severe; the most serious ones manifesting in the devastating Lesch-Nyhan syndrome, characterized by choreoathetosis or self-mutilation. Here we present a 30 years old male patient suffering from gout and mild psycho-motor impairment without Lesch Nyhan disease despite severe HPRT deficiency residual activity 0.02% with hypoxanthine, no activity at all with guanine as a substrate. The Curto's theory that neurologic impairment is dependent on VGPRT/VHPRT ratio is not confirmed by our observations. The finding of such a severe HPRT deficiency in a non-Lesch-Nyhan patient needs further investigation. G6PD deficiency was also referred together with beta-thalassemic trait. We have studied purine and pyridine nucleotide metabolism in the erythrocytes and discussed the literature. The bone marrow sample shows a megaloblastyc aspect.     

Efficacy and safety of allopurinol in patients with hypoxanthine-guanine phosphoribosyltransferase deficiency

Hypoxanthine-guanine phosphoribosyltransferase (HPRT) deficiency is a genetic disease of purine metabolism resulting in uric acid overproduction. Allopurinol, which inhibits the enzyme xanthine oxidase and reduces uric acid synthesis, is widely used for the treatment of gout and uric acid overproduction. The aim of the study was to analyze the long-term efficacy and safety of allopurinol in patients with HPRT deficiency. Nineteen patients (13 with Lesch-Nyhan syndrome and 6 with partial HPRT deficiency) were treated with allopurinol (mean dose, 6.4 mg/kg body weight per day; range, 3.7-9.7 mg/kg body weight per day) and followed up for at least 12 months (mean follow-up, 7.6 years). The efficacy of allopurinol was evaluated by serial measurement of purine metabolic parameters and renal function as well as by clinical manifestations. Safety was assessed by recording adverse events. Treatment with allopurinol normalized serum urate level in all patients and resulted in a mean reduction in serum urate of 47%. Allopurinol treatment was associated with a mean 74% reduction in urinary uric acid-to-creatinine ratio. In contrast, allopurinol treatment increased mean hypoxanthine and xanthine urinary excretion rates 5.4- and 9.5-fold, respectively, compared with baseline levels. The decrease in uric acid excretion in complete and partial HPRT-deficient patients was not accompanied by a stoichiometric substitution of hypoxanthine and xanthine excretion rates. Allopurinol-related biochemical changes were similar in patients with either complete or partial HPRT deficiency. Renal function remained stable or improved with treatment. Three patients had urolithiasis during allopurinol treatment. In 2 patients, xanthine stones were documented and they required allopurinol dose adjustments aimed at reducing excessive oxypurine excretion rates. No allopurinol hypersensitivity reactions occurred. Neurologic manifestations were not influenced by allopurinol therapy. In conclusion, allopurinol is efficacious and generally safe for the treatment of uric acid overproduction in patients with HPRT deficiencies. Xanthine lithiasis, developing as a consequence of allopurinol therapy, should be preventable by adjustment of allopurinol dose.